PROFORMA FOR SUBMISSION OF ANNUAL PROGRESS REPORT OFRESEARCH PROJECTS

600 Project Code6001 Institute Code No.6002 ICAR Code no.

IGFRI CI 2.9

Part-I: General Information

601 Name of Institute and Division6011 Name & Address oflnstitute: IGFRI, Jhansi-2840036012 Name of Division/Section : Crop Improvement Division6013 Location of Project : Jhansi

602 Project Title: Genetic improvement of pearl millet (bajra) and napier-bajra hybrids forhigher biomass production"

603 Priority Area:6031 Research Approach.: 02 (Basic research) and

03 (Process/ Technology development)

Applied res. Basic Res. Process/TechnologyDevelopment

01~ 02 03

Transfer ofTechnology

04

Specific Area:Genetic improvement of pearl millet (bajra) and napier-bajra hybrids for

higher biomass production"

605 Duration6051 Date of start: 20106052 Likely date of completion: 20156053 Period for which report submitted: 2012-13

606 Total cost of the Project:6061 Expenditure to date: Not applicable

607 Summary achievements: Please see annexure A.

608 Key words: Pearl millet, Napier grass, germplasm, hybrids, Interspecific hybrids,triploid

t-P~G__

610 Principal Investigator:6101 Name : Kumar Durgesh6102 Designation : Scientist (Plant Breeding)6103 Division : Crop Improvement Division6104 Location : Jhansi6105 Institute : IGFRI, Jhansi - 284003

612 Co- investigator:6121 Name : Dr. A.K Mishra6122 Designation : Principal Scientist6123 Division : PAR Division6124 Location : Jhansi6125 Institute : IGFRI, Jhansi - 284003

613 Co-investigator:6111 Name6112 Designation6113 Division6114 Location6115 Institute

611 Co-investigator:6111 Name6112 Designation6113 Division6114 Location6115 Institute

/Part - II: Investigator Profile

: Dr. C.K Gupta: Scientist: Seed Technology Division: Jhansi: IGFRI, Jhansi - 284003

: Manjunatha, N: Scientist: Seed Technology Division: Jhansi: IGFRI, Jhansi - 284003

Part - III: Technical Details

620 Introduction and objectives:6201 Immediate objectives:

~ Germplasm enrichment and characterisation of bajra (pearl millet) and napier.~ Generation of high yielding NB hybrids and bajra (pearl millet) hybrids

with high nutritional values.~ Cytologenetical studies in NB at enhanced ploidy levels for seed

producing NB.6202 Long term objectives:

~ Enhancing quality forage production by developing bajra (pearl millet)hybrids and NB hybrids.

621 Project Technical Profile:6211 Technical Programme :( indicate briefly plan of procedure, techniques,

instruments and special materials, organisms, special environment etc.)~ Acquisition from national international/institutes~ Multiplication of germplasm lines~ Identification and selection of parents and development of inbreds and

MS lines for hybrid production.~ Optimisation of pearl millet and P. purpureum and NB hybrid cytology~ Physiological characterisation of selected parents and the hybrids in

Pearl millet and NB hybrids~ Inducing polyploidy in NB using colchicine~ Development of hybrids in Pearl millet~ Hybrid development in NB.

6212 Man months involved of component project workers for the specified year

Dr. Kumar Durgesh 7 months

Dr. C.K Gupta 2 months

Dr. A.K Mishra 2 months

Manjunatha, N 1 months

Name of investigator Discipline Signature

Kumar Durgesh Plant Breeding ~ (f,/7ji)

Chandan Kumar Gupta Plant PhysiologyC~lt.~7,{t

A.K. Mishra Plant and Animal ~ 1-Relationship ~

Manjunatha, N Plant Pathology~~

Signature & Comments of the Head of the Division/Section

Signature & Comments of the Director

Annexure A

CI 2.9 Genetic improvement of pearl millet (bajra) and napier-bajra hybrids for higherbiomass production"

Collection/procurement of germ plasm

Germplasm procured: (Table-l )

1. Twenty seven lines of napier grass (Pennisetum purpureum) were imported/procuredfrom international agency-ILRI, Ethiopia.

2. And twenty two pearl millet lines were procured from USDA alongwith c:ne napiergrass line.

3. Thirty eight recombinant lines were also procured from ICRISAT targeting droughtand other fodder related traits.

4. Thirty five lines selected during Pearl millet scientist field day were procured fromICRISAT, Hyderabad which includes diverse lines .along with lines resistance tobiotic stresses.

Table-L Germplasm enrichment and multiplication

Crop Gemplasm/segregating Source Remarks No.lines

Napier Germplasm lines ILRI, Ethiopia - 27

Napier Germplasm lines USDA, USA - I

Pearl millet Germplasm lines . USDA, USA - 22

Pearl millet (RILs/ Germplasm ICRISAT, Drought and 38lines) Hyderabad other fodder

:i trait

Pearl millet Germplasm/MS lines ICRISAT, Biotic and 35Hyderabad abiotic stress

Total - - - 123lines

Germplasm multiplicationPearl millet:

About 273 lines were grown for multiplication with 3 checks (repeated)-Raj BajraChari, Giant Bajra Chari, and AVKB-19 mostly in paired rows (according to availability ofthe seeds). Data (Table-2) were taken from seventy three lines of pearl millet. Germplasmlines 5555, 2670, 8111 were topped in plant height (3.53m, 3.21 m, 3.18m), for tiller no.-germplasm no. 123, 10077 and 6140 were the best performer (8.6, 7.8 and 7.2), for no. ofleaves -germplasm lines 10077, 123 and 8111 were topped (49,34 and 31), for Green fodder

yield (GFY) -germplasm lines 123, 10077 and 6140 were best performer (1.7 kg,3.75g and2.7 g) and in brix % germplasm lines 19586, S13 and 2670 were best (21%,21%and 19 %) (Table-2). Selfed seeds of each lines were harvested and being thrashed. Fixationof panicle/spike for cytotogenetical study was also done.

Table-2. Five best performing germ plasm lines in following morphological traits.Characters Germplasm lines

PI. height 5555,2670,8111, 123, 13150

No. of tillers 123,10077,6140,8111,15536

No. of leaves 10077, 123,8111, Raj Bajra, 171

GFY 123,10077,6140,8111,15536

OM 6202, 04244, 7, 6863,

Brix% 19586, S13,2670,AVKB-19, 13016

Nutritional parameters and physiological and biochemical screening of Pearl milletIines/ germ plasm

Newly procured accessions of pearl millet along with last year selected lines generationmaterial were characterized for nutritional and biochemical! physiological parameters. Sixtynine lines of pearl millet were screened for quality parameters viz. crude protein, lignincontent, NOF%, and cellulose content etc. which is represented in (Table-3). Forphysiological/enzymatic screening leaf samples were extracted.

Table-3: Selected lines on the basis of better nutritive value(s) over checks (mean)

Characters Selected lines

Crude Protein PN4-16, PN5-15, PN3-44, PN3-46, PN4-6

NOF (%) PN4-2, PN5-l3, PN3-l4, PNI-47, PN3-46

Lignin(%) P 3-16, PN 1-63,PN3-62, P 3-4, PN5-5

Cellulose PN5-1, PN3-46, PNl-24, PNI-47, PNI-67, PNl-65

Protein estimation and enzyme assayPresence of antioxidative enzymes is an important indicator of plant health. Catalase (CAT)and peroxidase (POX) are antioxidative enzymes which are instrumental in degrading H202

We estimated CAT and POX activity in selected lines at 50% flowering stage. All theselected lines under study showed good activity of both the enzymes which indicate theirbetter potential for tolerance towards extreme environmental conditions. Data of twelve linesare tabulated here (Table 4).Total soluble proteins were also estimated by Bradford methods.

Table-4. Total soluble protein, CAT and POX activity in selected lines of pearl millet.

Lines Protein (ug/g FW) CA T (Unit/mg protein) POX (Unit/mg protein)

CK39 7.54 0.032 0.04

CK33 3.96 0.039 0.045

CK 32 9.82 0.03 0.041 ICK41 7.70 0.028 0.038

CK39 8.18 0.035 0.039

CK40 8.94 0.033 0.04

CK37 4.02 0.03 0.04

CK38 7.76 0.03 0.04

CK40 8.60 0.032 0.042

CK41 6.76 0.032 0.041

CK35 6.15 0.03 0.043

CK31 6.81 0.03 0.045

Selection and hybridisation:About 30 better performing lines were selected in Pearl millet and hybridisation wereperformed for the same line with napier grass as male and pearl millet as female parent forthe optimisation of interspecific hybridisation and putative NB hybrids were made. For MSlines development hybridisation were done between forage type pearl millet and Ascytoplasm along with bmr mutant.

Screening of Pearl millet for Disease resistance: At IGFRI, in pearlmillet field experimentplot about 120 germplasm lines were observed for foliar diseases and recorded severity ofrust, Pyricylaria leaf spot and Cercospora leaf spot. The severity was ranged from 5% to

40%. The diseased samples were collected and etiology was confirmed through microscopicexamination (Fig. 1) About 20 resistant lines were selected which are potential genotypes andcan be used In resistant breeding programme.

r1

Figure-I. Microscopic observation of foliar disease pathogens such as Puccinia striata,Pyricularia grisea and Cercospora pennisiti respectively.

Interspecific hybridisation/ploidy enhancement:

Trispecific hybrid (TSH) of pearl millet was developed from crosses (out of more than 45crosses) between pearl millet. Pennisetum squamulatum and P. purpureum napier grass.Preliminary investigation on relative DNA content proves the trispecific nature of thehybrids. This was proved by flow cytometric data on relative DNA content (fig.-2). Ploidyenhancements of 3X NB hybrids were also attempted by colchicine and it requires furtherrefinement to get higher level of polyploidy in NB hybrid.

1gg

2 TSH

gg

o 50 100 150 200 2E 0 50 100 150 200 2!

Fig. 2 Relative DNA content of TSH hybrid as shown in flow cytometry data.

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INDIAN COUNCIL OF AGRICULTURAL RESEARCH

RESEARCH PROJECT PROFORMA FOR MONITORING ANNUAL PROGRESS (RPF-II)

c-\ ~ ~1. Institute Project Code: IGFRICI2.9

2. Project Title: : "Genetic improvement of pearl millet (bajra) and napier-bajra hybrids

for higher biomass production"

3. Reporting Period :2012-13

4. Project Duration: Date of Start - 2011 Likely Date of Completion -2015

5. Project Team (Name(s) and designation of PI,CC-PIand all project Co-Pis, (with time spent forthe project) if any additions/deletions

I

S. Name, designation and Status in the Time to" be Work components assigned toNo. institute project (PI/CC-PI/ spent{%) individual scientist

Co-PI)1 PI 55 Acquisition from national

Dr. Kumar Durgesh, international/institutesScientist, IGFRI Multiplication of germplasm

lines

Identification and selection ofparents and development ofinbreds and MS lines for hybridproduction.Optimisation of pearl milletand P. purpureum and NBhybrid cytologyInducing polyploidy in NBusing colchicineDevelopment of hybrids inPearl milletHybrid development in NB.

2-. Dr. A K Mishra Co-PI 15. Test of quality parameters

3 Dr. Chandan K. Gupta, Co-PI 15 PhysiologicalScientist, IGFRI characterisation of selected

parents and the hybrids inPearl millet and NB hybrids

4- Manjunatha, N Co-PI 15 Pathological screening of pearlmillet/napier lines

6. (a) Activities and outputs earmarked for the year (as per activities schedule given in RPF-I)

Objective wise Activity .Scientist % of activity % achievedresponsible envisaged to be as targeted

completed asper RPP-I

l.Germplasm SeeAnnexure 1 KDurgesh 100% 100%

enrichment and Dr. ChandanK To be completed -

characterization Gupta in this week01 Manjunatha, N 100% 100%

of bajra andnapier lines

2. Cytogenetical SeeAnnexure 1 K Durgesh 100%and 100%Studies in NB contd.,

hybrids and theirpolyploidsGeneration of SeeAnnexure 1 KDurgesh 100%high yielding Dr.A k Mishra 100%

with betternutritive valueBajra (PearlMillet) and NBhybrids

(b) If shortfall/addition, reasons for the same and how to catch up with the intended activities

7. Annual Progress Report (research results and achievements in bullets) : Annexure- A

8. Output During Period Under Report: Annexure- A

a. Special attainments/innovationsb. List of Publications (one copy each to be submitted with RPP-II)

i. Research papersii. Reports/Manuals

iii. Working and Concept Papersiv. Popular articles

v.. Books/Book Chaptersvi. Extension Bulletins

c. Intellectual Property Generation: : See Annexure -A(Patents - filed/obtained; Copyrights- filed/obtained; Designs- filed/obtained;Registration details of variety/germplasm/accession if any)

d. Presentation in Workshop/Seminars/Symposia/Conferencese. (relevant to the project in which scientists have participated)f. . Details of technology developed: : See Annexure -A

(Crop-based; Animal-based, including vaccines; Biological- biofertilizer,biopesticide, etc; IT based - database, software; Any other - please specify)

g. Trainings/demonstrations organizedh. Training received

i. Any other relevant information

9. Constraints experienced, if any

10. LessonsLearnt

11. Evaluation

(a) Self evaluation of the project for the period under report by the PIwith rating Din the scale of 1 to 10

(b) Evaluation by Pion the contribution of the team in the project including self

S. Name Status in the project Rating in the scale of 1 to 10No. (PI/CC-PI/Co-PI)

D12. Signature of PI, CC-PI(s),all Co-Pis

13. Signature (with specific comments on progress/achievements, shortfall andconstraints along with rating ofthe project in the scale of 1 to 10) of DHead of Division/Regional Center / Section

14. Comments of IRC

15. Signature (with specific comments on progress/achievements, shortfall Dand constraints along with rating of the project in the scale of 1 to 10)of JD (R)/ Director

Annexure A

CI 2.9 Genetic improvement of pearl millet (bajra) and napier-bajra hybrids forhigher biomass production"

Collection/procurement of germ plasm

Germplasm procured:

1. Twenty seven lines of napier grass (Pennisetum purpureum) were

imported/procured from international agency-ILRI, Ethiopia which weresown in pot.

2. And twenty two pearl millet lines were procured from USDA alongwith onenapier grass line.

3. Thirty eight recombinant lines were also procured from ICRISAT targetingdrought and other fodder related traits.

4. Thirty five lines selected during Pearl millet scientist field day wereprocured from ICRISAT, Hyderabad which includes diverse lines along withlines resistance to biotic stresses.

Germplasm multiplication

Pearl millet:

About 273 lines were grown for multiplication with 3 checks (repeated)-Raj

Bajra Chari, Giant Bajra Chari, and AVKB-19 mostly in paired rows (according to

availability of the seeds); Data (Table-l) were taken from seventy three lines of

pearl millet. Germplasm lines 5555, 2670, 8111 were topped in plant height

(3.53m, 3.21m, 3.18m), for tiller no.- germ plasm no. 123, 10077 and 6140 were

the best performer (8.6, 7.8 and 7.2), for no. of leaves -gerrnplasm lines 10077, 123

and 8111 were topped (49,34 and 31), for Green fodder yield (GFY) -germ plasm

lines 123, 10077 and 6140 were best performer (1.7 kg,3.75g and2.7 g) and in brix

% germ plasm lines 19586, S13 and 2670 were best (21%, 21% and 19 %). Selfed

seeds of each lines were harvested and being thrashed. Fixation of panicle/spike

for cytotogenetical study was also done.

Characterization and multiplication Pearl millet lines/ germ plasm

All newly procured accessions of pearl millet along with last year selected

lines F 6/7 generation material were characterize for morphological traits viz., plant

ht., no. of tillrers, , no. of leaves, GFY, Brix % etc. Ten best performing germ plasm

lines are in following morphological traits (Table-l).

Table-1. Ten best performing germplasm lines in following morphological traits.PI. height No. of tillers No. of leaves GFY Brix %

Germplasm lines/Check5555 123 10077 123 195862670 10077 123 10077 s138111 6140 8111 6140 2670123 8111 c3 8111 cl13150 15536 171 15536 130162045 5222 4155 5222 composite13016 2045 146 2045 cl19628 13150 467 13150 c34232 171 5555 171 s1282 6289 1229. 6289 82

Selection and hybridisation: some about 30 better performing lines were selectedin Pearl millet and hybridisation were performed for the same line with napiergrass as male and pearl millet as female parent for the optimisation of interspecifichybridisation and putative NB hybrids were made. For MS lines developmenthybridisation were done between forage type pearl millet and As cytoplasmalongwith bmr mutant.Screening of Pearl millet for Disease resistance: At IGFRI, in pearlmillet fieldexperiment plot about 120 germplasm lines were observed for foliar diseasesandrecorded severity of rust, Pyricylaria leaf spot and Cercospora leaf spot. Theseverity was ranged from 5% to 40%. The diseased samples were collected andetiology was confirmed through microscopic examination (Fig. 1) About 20resistant lines were selected which are potential genotypes and can be used inresistant breeding programme.

Figure-1. Microscopic observation of foliar disease pathogens such as Puccinia

striata, Pyricularia grisea and Cercospora pennisiti respectively.

-...:...---------' ,~' ~,

Nutritional parameters and physiological and biochemical screening:

Sixty nine lines of pearl millet were screened for quality parameters viz. ash %,lignin content, 72% H2S04, NDF%, OM%, Cellulose etc. which is represented in(Table-2). For physiological/enzymatic screening leaf samples were extracted.Interspecific hybridisation/ploidy enhancement:

Trispecific hybrid (TSH) of pearl millet was developed from crosses (out of morethan 45 crosses) between pearl millet. Pennisetum squamulatum and P. purpureum

napier grass. Preliminary investigation on relative DNA content proves thetrispecific nature of the hybrids. Which was proved by flow cytometric data onrelative DNA content(fig.-2). Ploidy enhancement of 3X NB hybrids were alsoattempted by colchicine.

1gg

2 TSH

gg

o 50 100 150 200 250 50 100 150 200 2!

Fig. 2 Relative DNA content of TSHhybrid as shown in flow cytometry data.

Details of Training/ Refresher Course/Summer/Winter Institutes/Seminars/ : No

Conferences/ Symposia/ Workshops attended within India and on Deputationabroad:Visit:I attended national workshop on forage crop (Kharif) at Pune where I got All India

Co-ordinated trial of BN hybrids for IGFRI, Jhansi station.

IlH177 a cow pea variety was identified for the release by varieties Identification

committee at All India Coordinated Research Project on Forage Crops in Pune

national workshop in which I am associated.

I attended 3-day scientist field day for pearl millet at ICRISAT, Hyderabad

Tc4t'1--- Statistical representation of nutritive value of selected 69 forage tvpeslil1g_~earl millet:

72% H2SO4 Cellulose Lignin % NDF% Ash % OM%Statistical Parameters ADF%

45.59 33.28 5.51 62.22 11.09 11.09Mean 45.58

5.54 2.90 0.15 0.42 0.21 0.21Standard Error 5.541959

39.86 30.82 5.54 62.00 11.00 11.00Median 39.86

43.00 30.00 3.86 62.00 10.00 10.00Mode 43

46.03 24.11 1.28 3.53 1.74 1.74Standard Deviation 46.03

2119.22 581.16 1.64 12.44 3;02 3.02Sample Variance 2119.21

68.51 64.63 5.38 1.02 -0.55 -0.55Kurtosis 68.51

8.26 7.89 1.54 -0.07 0.00 0.00Skewness 8.26

386.49 225.53 8.30 20.00 7.00 7.00Range 386.49

35.31 2.00 2.91 51.00 7.00 7.00Minimum 35.31

421.80 227.53 11.21 71.00 14.00 14.00Maximum 421.8

\3145.38 2296.10 380.14 4293.24· 764.94 764.94Sum 3145.38

69.00 69.00 69.00 69.00 69.00 69.00Count 69

1.68 5.79 0.31 0.85 0.42 0.42Confidence Level (95.0%) 11.05